Neurotoxicity of mycotoxins produced in vitro by Penicillium roqueforti isolated from maize and grass silage

Fungal growth in human foods and animal feeds causes profound damage indicating a general spoilage, nutritional
losses, and the formation ofmycotoxins. Thirty apparently contaminated maize and grass silage samples were analyzed
for the presence of total fungi. Penicillium roqueforti were isolated from all (100%) moldy silage samples on
general and selective culture media. Furthermore, P. roqueforti-positive samples culture media subjected to the
toxin extraction and toxins of patulin, penicillic acid, mycophenolic acid, and roquefortin-C (ROQ-C) were identified
by means of high-performance liquid chromatography method. Cytotoxicity of identified toxins was investigated
on neuro-2a cells. Alamar blue reduction, neutral red uptake, and intracellular adenosine triphosphate (ATP)
content assays indicated that patulin and ROQ-C exert the strongest and weakest toxicity, respectively. Reactive
oxygen species (ROS) generation by the toxins-exposed cells was measured, and the results supported the
mitochondrial and lysosomal dysfunction and ATP depletion in exposed cells. Our data suggest that P. roqueforti
is the widely present mold in analyzed maize and grass silage samples, which is able to produce toxins that
cause neurotoxicity. This finding may explain in part some neuronal disorders in animals, which are fed contaminated
feedstuffs with mentioned fungus. Moreover, mitochondrial and lysosomal dysfunction, intracellular
ATP depletion, and the excessive ROS generation were found as the mechanisms of cytotoxicity for P. roqueforti-
produced toxins